Cholestane is a fully saturated alkane with a structure analogous to cholesterol. We observed fluorescence emission from cholestane with a maximum near 235 nm when excited with picosecond laser pulses at 298 nm. This emission at 235 nm was shown to be due to two-photon excitation. The fluorescence lifetime of cholestane in cyclopentane was found to be near 1.4 ns. Cholestane is efficiently collisionaly quenched by methanol and oxygen. These results suggest that cholestane or its analogs can be used as intrinsic probes in biological membranes. Two-photon excitation avoids the use of vacuum ultraviolet wavelength (130-170 nm), which require special optics and exclusion of oxygen. This observation may lead to the more general use of alkane fluorescence as a probe of cell membranes.